白细胞介素-33在肺癌细胞恶性生长中的作用及机制的基础研究

发布时间：2018-03-22 06:14

本文选题：肺癌　切入点：非小细胞肺癌　出处：《青岛大学》2017年硕士论文　论文类型：学位论文

【摘要】：目的:肺癌是目前全球恶性肿瘤累及死亡的首要原因,发病率高,临床缺乏有效治疗手段,死亡率高,是严重危害人群健康的一大难题。恶性生长是肺癌细胞的重要生物学特征,也是评价临床治疗效果的关键指标,然而肺癌细胞恶性生长的维持和促进因素仍不清楚。深入研究肺癌细胞恶性生长的维持和促进因素,对理解肺癌发病机制和研发临床治疗新策略有重要意义。本课题以肺癌患者肿瘤细胞为靶点,探讨白细胞介素-33(IL-33)在肺癌细胞恶性生长中的作用及其可能机制。方法:肿瘤细胞分离采用肿瘤细胞分离试剂盒,细胞转染用Lonza转染试剂盒,转染效率通过qPCR和流式细胞术检测。肿瘤细胞体外生长用MTT法检测,肿瘤细胞体内生长情况用人源化小鼠模型分析。IL-33、ST2和葡萄糖转运蛋白1(GLUT1)的表达水平用qPCR、免疫组化染色或流式细胞术检测,葡萄糖摄取率和乳酸产生水平分别按照试剂盒说明书进行。结果:IL-33与其受体ST2在肿瘤组织中的表达水平显著高于在癌旁组织中的表达水平;IL-33和ST2的表达水平与肺癌肿瘤分期显著正相关,在晚期肺癌患者体内存在更高水平的表达;IL-33和ST2表达水平可提示肿瘤细胞的低分化程度,在低分化肺癌细胞中存在显著高表达。通过转染人IL-33正义表达载体上调肺癌细胞内IL-33表达水平可在体外和体内有效促进肿瘤细胞的恶性生长,通过转染人IL-33 shRNA下调肺癌细胞内IL-33表达水平可明显抑制肺癌细胞在体外和体内的恶性生长能力。IL-33蛋白刺激肺癌细胞可以直接促进肺癌细胞在体外和体内的恶性生长能力,上调ST2表达水平可进一步增强IL-33对肺癌细胞恶性生长的促进作用,下调ST2表达水平可阻断IL-33对肺癌细胞恶性生长的促进作用,ST2中和抗体亦有效抑制IL-33对肺癌细胞恶性生长的影响。IL-33影响肺癌细胞恶性生长的分子机制在于IL-33蛋白刺激肺癌细胞能够上调肿瘤细胞膜GLUT1蛋白的水平,该过程可被ST2中和性抗体所阻断;IL-33蛋白亦可增强肺癌细胞的葡萄糖摄取能力和乳酸产生水平;基因敲除GLUT1在肺癌细胞中的表达可显著抑制IL-33对肺癌细胞恶性生长的促进作用。结论:IL-33是临床肺癌患者肿瘤进展中的重要促癌因子,IL-33能够维持和促进肺癌细胞的恶性生长能力。IL-33维持和促进肺癌细胞恶性生长可以通过ST2受体识别实现,下调或阻断IL-33/ST2信号可显著抑制肺癌细胞的恶性生长。IL-33/ST2信号能够增强肺癌细胞膜GLUT1蛋白水平进而促进肺癌细胞的葡萄糖摄取能力和糖酵解,为肺癌患者肿瘤细胞恶性生长提供能量支持。基于IL-33/ST2信号的干扰策略可能是临床控制肺癌细胞恶性生长的有效手段。该研究不仅有助于深入理解临床肺癌发生发展的分子机制,更可为研发临床治疗肺癌新策略提供理论依据,具有重要的理论意义和现实意义。
[Abstract]:Objective: lung cancer is the leading cause of death from malignant tumors in the world at present. The incidence of lung cancer is high, the clinical treatment is lack of effective treatment, and the mortality rate is high. Malignant growth is an important biological feature of lung cancer cells and a key index to evaluate the clinical therapeutic effect. However, the maintenance and promotion factors of malignant growth of lung cancer cells are still unclear. It is of great significance to understand the pathogenesis of lung cancer and to develop new clinical treatment strategies. To explore the role of interleukin-33 (IL-33) in the malignant growth of lung cancer cells and its possible mechanism. Methods: tumor cells were separated by tumor cell separation kit and Lonza transfection kit was used for cell transfection. The transfection efficiency was detected by qPCR and flow cytometry. The growth of tumor cells in vitro was detected by MTT. In vivo growth of tumor cells was detected by qPCR, immunohistochemical staining or flow cytometry to analyze the expression of. IL-33 / ST2 and glucose transporter 1 (GLUT1) using a humanized mouse model. The glucose uptake rate and lactic acid production level were carried out in accordance with the kit instructions. Results the expression levels of ST2 and ST2 in tumor tissues were significantly higher than those in paracancerous tissues. The expression levels of IL-33 and ST2 in lung and lung were significantly higher than those in paracancerous tissues. There was a significant positive correlation between tumor staging and tumor staging. The higher expression of IL-33 and ST2 in patients with advanced lung cancer may indicate the low differentiation of tumor cells. The expression of IL-33 in lung cancer cells was up-regulated by transfection of human IL-33 sense expression vector, which could effectively promote the malignant growth of tumor cells in vitro and in vivo. Down-regulating the expression of IL-33 in lung cancer cells by transfection of human IL-33 shRNA could significantly inhibit the malignant growth ability of lung cancer cells in vitro and in vivo. IL-33 protein stimulated lung cancer cells can directly promote the malignant growth ability of lung cancer cells in vitro and in vivo. Upregulating the expression of ST2 can further enhance the role of IL-33 in promoting the malignant growth of lung cancer cells. Down-regulation of ST2 expression can block the effect of IL-33 on the malignant growth of lung cancer cells. ST2 neutralizing antibody can also effectively inhibit the effect of IL-33 on the malignant growth of lung cancer cells. The molecular mechanism of IL-33 affecting the malignant growth of lung cancer cells lies in the IL-33 protein prick. Lung cancer cells can up-regulate the level of GLUT1 protein in tumor cell membrane. This process can be blocked by ST2 neutralizing antibody. IL-33 protein can also enhance glucose uptake and lactic acid production in lung cancer cells. The expression of knockout GLUT1 in lung cancer cells can significantly inhibit the role of IL-33 in promoting the malignant growth of lung cancer cells. Conclusion: IL-33 is an important tumor promoting factor in clinical lung cancer patients, which can maintain and promote the growth of lung cancer cells. The ability of malignant growth. IL-33 to maintain and promote the malignant growth of lung cancer cells can be achieved by ST2 receptor recognition. Down-regulating or blocking the IL-33/ST2 signal can significantly inhibit the malignant growth of lung cancer cells. IL-33 / ST2 signal can enhance the GLUT1 protein level of lung cancer cell membrane and promote glucose uptake and glycolysis of lung cancer cells. The interference strategy based on IL-33/ST2 signal may be an effective method to control the malignant growth of lung cancer cells. This study is not only helpful to understand the molecular mechanism of the occurrence and development of clinical lung cancer. It can provide a theoretical basis for the development of new strategies for the treatment of lung cancer, and has important theoretical and practical significance.
【学位授予单位】：青岛大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：R734.2

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